利用VIGS技术研究棉花GhPPO1基因的抗虫作用

发布时间：2018-01-23 22:25

  本文关键词： 棉花 病毒介导的基因沉默(VIGS) 多酚氧化酶基因(PPO) 抗虫性 基因表达　出处：《农业生物技术学报》2017年05期 　论文类型：期刊论文

【摘要】：多酚氧化酶(polyphenol oxidase,PPO)是一类含铜的氧化还原酶,能够抵御植食性昆虫或病原菌的危害。为了研究棉花(Gossypium arboretum)中PPO1基因的抗虫作用,本研究利用病毒介导的基因沉默(virus-induced gene silencing,VIGS),构建基因沉默重组载体pTRV2-GhPPO1,转化农杆菌(Agrobacterium tumefaciens)GV3101,并侵染子叶展平的棉花植株;通过qRT-PCR检测沉默效果,并用获得的棉花植株饲喂二龄初期棉铃虫(Helicoverpa armigera)幼虫,对沉默后棉花植株的抗虫性及GhPPO1基因表达量的变化进行检测。结果表明,利用烟草脆裂病毒(Tobacco rattle virus,TRV)体系成功抑制了棉花GhPPO1的表达,沉默后GhPPO1基因相对表达量仅为对照组(空载体pTRV0)表达量的20%(P0.05)。与饲喂正常棉花叶片的棉铃虫相比,饲喂GhPPO1基因沉默后棉花叶片的棉铃虫72 h后,体重增加86%,表明GhPPO1是棉花抗虫物质中的一类防御机制基因。棉铃虫直接诱导18、72 h的叶片中GhPPO1的表达量高于间接诱导叶片,证明沉默该基因后,消弱了棉花植株直接防御的能力,致使棉铃虫增重效果显著。由此推测该基因可能参与棉花直接防御反应。本研究对于GhPPO1在棉花防御植食性昆虫中的作用具有重要意义,为棉铃虫与棉花诱导防御之间的关系提供基础资料。
[Abstract]:Polyphenol oxidase (PPO) is a kind of copper-containing redox enzyme. In order to study the insect-resistance of PPO1 gene in cotton Gossypium arboretum. In this study, virus-induced gene silencing (VIGSs) was used in virus-mediated gene silencing. The recombinant vector pTRV2-GhPPO1 was constructed and transformed into Agrobacterium tumefaciens)GV3101. And infecting cotton plants with flattened cotyledons; The silencing effect was detected by qRT-PCR, and the obtained cotton plants were fed with Helicoverpa armigera larvae. The insecticidal resistance and the expression of GhPPO1 gene in cotton plants after silencing were detected. The results showed that tobacco rattle virus was used for tobacco embrittlement virus. TRV) system successfully inhibited the expression of GhPPO1 in cotton. The relative expression of GhPPO1 gene after silencing was only 20% of that of the control group (empty vector pTRV0), compared with that of cotton bollworm fed normal cotton leaves. Cotton bollworm (Helicoverpa armigera) fed with GhPPO1 gene silencing for 72 h increased its weight by 86%. The results showed that GhPPO1 was a kind of defense gene in cotton insect-resistant materials, and the expression of GhPPO1 in leaves induced by Helicoverpa armigera for 1872 h was higher than that in indirectly induced leaves. It was proved that silencing the gene weakened the ability of cotton plants to defend directly. It is suggested that this gene may be involved in direct defense response of cotton. This study is of great significance for the role of GhPPO1 in cotton plant insect defense. To provide basic data for the relationship between cotton bollworm and cotton induced defense.
【作者单位】： 新疆农业大学农学院;中国农业科学院棉花研究所/棉花生物学国家重点试验室;
【基金】：国家转基因生物新品种培育科技重大专项(No.2016ZX08012-004)
【分类号】：S435.62
【正文快照】： 棉花(Gossypium arboreum)是重要的经济作物,是纺织纤维、饲料等的重要来源。随着棉花全基因组测序的完成,应用分子工具和技术资源分析基因功能至关重要(Wang et al.,2012)。棉花的生长周期长、基因组功能冗杂,遗传转化过程费力耗时并且通常很难实现(Gu et al.,2014)。病毒介，

本文编号：1458388